title: Detection and quantification of specific DNA adducts by liquid chromatography-tandem mass spectrometry in the livers of rats given estragole at the carcinogenic dose.
abstract: Estragole (ES) is a natural constituent of several herbs and spices that acts as a carcinogen in the livers of rodents. Given that the proximal electrophilic form of ES with a reactive carbocation is generated by cytochrome P450 and a sulfotransferase metabolizing pathway, there is a possibility that the resultant covalent adducts with DNA bases may play a key role in carcinogenesis. The existence of ES-specific deoxyguanosine (dG) and deoxyadenosine (dA) adducts has already been reported with the precise chemical structures of the dG adducts being confirmed. In the present study, we examined ES-specific dA adduct formation using LC-ESI/MS after the reaction of dA with 1'-acetoxy-ES produced by a sulfotransferase metabolic pathway mimic. Although two peaks were observed in the LC-ESI/MS chromatogram, the identification of ES-3'-N(6)-dA as the measurable peak was determined by NMR analysis. To confirm ES-specific dG and dA adduct formation in vivo, an isotope dilution LC-ESI/MS/MS method applicable to in vivo samples for ES-3'-N(6)-dA together with the two major dG adducts, that is, ES-3'-C8-dG and ES-3'-N(2)-dG, was developed using selected ion recording. The limit of quantification was 0.2 fmol on column for ES-3'-C8-dG and ES-3'-N(2)-dG and 0.06 fmol on column for ES-3'-N(6)-dA, respectively. Using the developing analytical method, we attempted to measure adduct levels in the livers of rats treated with ES at a possible carcinogenic dose (600 mg/kg bw) for 4 weeks. ES-3'-C8-dG, ES-N(2)-dG, and ES-3'-N(6)-dA were detected at levels of 3.5 ± 0.4, 4.8 ± 0.8, and 20.5 ± 1.6/10(6) dG or dA in the livers of ES-treated rats. This quantitative data and newly developed technique for adduct observation in vivo might be helpful for ES hepatocarcinogenesis investigations.
authors.affiliation: Division of Pathology, National Institute of Health Sciences , 1-18-1 Kamiyoga, Setagaya-ku, Tokyo, Japan.
authors.forename: Yuji
authors.initials: Y
authors.lastname: Ishii
authors.forename: Yuta
authors.initials: Y
authors.lastname: Suzuki
authors.forename: Daisuke
authors.initials: D
authors.lastname: Hibi
authors.forename: Meilan
authors.initials: M
authors.lastname: Jin
authors.forename: Kiyoshi
authors.initials: K
authors.lastname: Fukuhara
authors.forename: Takashi
authors.initials: T
authors.lastname: Umemura
authors.forename: Akiyoshi
authors.initials: A
authors.lastname: Nishikawa
mesh_terms: D000081248:Allylbenzene Derivatives; D000818:Animals; D000840:Anisoles / Q000737:chemistry* / Q000633:toxicity; D002273:Carcinogens / Q000737:chemistry* / Q000633:toxicity; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D018736:DNA Adducts / Q000032:analysis* / Q000737:chemistry; D008099:Liver / Q000737:chemistry* / Q000187:drug effects / Q000378:metabolism; D008297:Male; D051381:Rats; D011916:Rats, Inbred F344; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D053719:Tandem Mass Spectrometry
chemical_list: D000081248:Allylbenzene Derivatives; D000840:Anisoles; D002273:Carcinogens; D018736:DNA Adducts; C007633:estragole